林洁, 张士煜, 吕琦, 赵洪彬, 李旋, 童亮, 马岚. 58型人乳头瘤病毒E7蛋白单克隆抗体的制备及鉴定[J]. 云南大学学报(自然科学版), 2012, 34(6): 731-737.
引用本文: 林洁, 张士煜, 吕琦, 赵洪彬, 李旋, 童亮, 马岚. 58型人乳头瘤病毒E7蛋白单克隆抗体的制备及鉴定[J]. 云南大学学报(自然科学版), 2012, 34(6): 731-737.
LIN Jie, ZHANG Shi-yi, LÜ Qi, ZHAO Hong-bin, LI Xuan, TONG Liang, MA Lan. Preparation and identification of monoclonal antibody against E7 oncoproteinfrom human papillomavirus type 58[J]. Journal of Yunnan University: Natural Sciences Edition, 2012, 34(6): 731-737.
Citation: LIN Jie, ZHANG Shi-yi, LÜ Qi, ZHAO Hong-bin, LI Xuan, TONG Liang, MA Lan. Preparation and identification of monoclonal antibody against E7 oncoproteinfrom human papillomavirus type 58[J]. Journal of Yunnan University: Natural Sciences Edition, 2012, 34(6): 731-737.

58型人乳头瘤病毒E7蛋白单克隆抗体的制备及鉴定

Preparation and identification of monoclonal antibody against E7 oncoproteinfrom human papillomavirus type 58

  • 摘要: 为获得抗58型人乳头瘤病毒(HPV58)E7蛋白的单克隆抗体,乳糖诱导重组工程菌表达HPV58 E7融合蛋白,以纯化后的该重组蛋白为免疫原,经细胞融合和有限稀释后,得到6株分泌抗HPV58 E7单克隆抗体(McAb)的工程细胞株,并采用体内接种方法及亲和柱层析纯化技术制备6株抗体.酶联免疫吸附测定(ELISA)、Western-blot实验和细胞免疫荧光检测结果表明,所得抗体均为IgG2a类,κ型,抗体效价均可达10-5,并且能够与原核及真核系统表达的HPV58 E7蛋白发生特异性抗原-抗体反应.该抗体的成功制备为后续HPV58 E7致癌生物学研究及相关肿瘤诊断试剂的开发奠定基础.

     

    Abstract: To obtain monoclonal antibodies (McAbs) against E7 oncoprotein of human papillomavirus type 58 (HPV-58),lactose induction was applied to express GST-HPV58E7 fusion protein in recombinant bacterial strain.Using simply GST affinity chromatographic separation,the target protein was purified and used as a novel antigen to produce antibodies.After mouse immunization,somatic hybridization,cloned culture,positive hybridoma scanning,ascites preparation,rProtein,and affinity purification,6 strains of purified and high titer anti-HPV58E7 McAbs were prepared.Further studies used ELISA,Western-blot and immunofluorescence,and proved these obtained antibodies could specifically react with HPV-58 E7 which was expressed either in prokaryotic or eukaryotic cells.Moreover,another indirect ELISA also revealed that these 6 strains of McAbs were IgG2a κ type antibodies and bound with the same epitope in entire sequence of HPV-58 E7 oncoprotein. In conclusion,the prepared McAbs provides the basis for tumor biology research and the development of related diagnostic reagents which are targeting HPV-58 E7,and these studies are very important to effective screening,diagnosis,prevention and treatment of cervical cancer caused by HR-HPV detect in our country.

     

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